The biological function of RNA is mediated by its structure. mRNA is generally thought of as a linear molecule which contains the information for directing protein synthesis within the sequence of ribonucleotides. Recently, studies have revealed a number of secondary and tertiary structures in mRNA which are important for its function (See; I. Tinoco, P. W. Davis, C. C. Hardin, J. D. Puglisi, G. T. Walker, Cold. Spring. Harb. Symp. Quant. Biol. 52, 135 (1987). Secondary structural elements in RNA are formed largely by Watson-Crick type interactions between different regions of the same RNA molecule. Important secondary structural elements include intramolecular double stranded regions, hairpin loops, bulges in duplex RNA and internal loops. Tertiary structural elements are formed when secondary structural elements come in contact with each other or with single stranded regions to produce a more complex, three dimensional structure.
Very little is known about the precise three dimensional structure of RNA. However, there have recently been a number of research efforts which have shown that RNA structures, including single stranded, secondary, and tertiary structures, have important biological functions beyond simply encoding information to make proteins in linear sequences. Some of these correlations have been discussed in: I. Tinoco, P. W. Davis, C. C. Hardin, J. D. Puglisi, G. T. Walker, Cold. Spring. Harb. Symp. Quant. Biol. 52, 135 (1987); 0. Resnekov, M. Kessler, Y. Aloni, J. Biol. Chem. 264, 9953 (1989); C. Tuerk, P. Gauss, C. Thermes, et al, Proc. Natl. Acad. Sci. U. S. A. 85, 1364 (1988); D. E. Larson, B. H. Sells, Mol. Cell. Biochem. 74, 5 (1987); and G. Knapp, Methods Enzymol. 180, 192 (1989).
Oligonucleotides have been evaluated for effect on HIV. Agarwal and coworkers have used oligonucleotide analogs targeted to the splice donor/acceptor site to inhibit HIV infection in early infected and chronically infected cells. S. Agarwal, T. Ikeuchi, D. Sun, P. S. Sarin, A. Konopka, J. Maizel, Proc. Natl. Acad. Sci. USA 86:7790 (1989). Sarin and coworkers have also used chemically modified oligonucleotide analogs targeted to the cap and splice donor/acceptor sites. P. S. Sarin, S. Agarwal, M. P. Civerira, J. Goodchild, T. Ikeuchi, P. C. Zamecnik, Proc. Natl. Acad. Sci. USA 85:7448 (1988). Zaia and coworkers have also used an oligonucleotide analog targeted to a splice acceptor site to inhibit HIV. Zaia, J. A., J. J. Rossi, G. J. Murakawa, P. A. Spallone, D. A. Stephens, B. E. Kaplan, J. Virol. 62:3914 (1988). Matsukura and coworkers have synthesized oligonucleotide analogs targeted to the initiation of translation of the rev gene mRNA. M. Matsukura, K. Shinozuka, G. Zon, et al., Proc Natl. Acad. Sci. USA, 84:7706 (1987); R. L. Letsinger, G. R. Zhang, D. K. Sun, T. Ikeuchi, P. S. Sarin, Proc. Natl. Acad. Sci. USA 86:6553 (1989). Mori and coworkers have used a different oligonucleotide analog targeted to the same region as Matsukura et al., K. Mori, C. Boiziau, C. Cazenave et al., Nucleic Acids Res. 17:8207 (1989). Shibahara and coworkers have used oligonucleotide analogs targeted to a splice acceptor site as well as the reverse transcriptase primer binding site. S. Shibahara, S. Mukai, H. Morisawa, H. Nakashima, S. Kobayashi, N. Yamamoto, Nucl. Acids Res. 17:239 (1989). Letsinger and coworkers have synthesized and tested oligonucleotide analogs with conjugated cholesterol targeted to a splice site. K. Mori, C. Boiziau, C. Cazenave, et al., Nucleic Acids Res. 17:8207 (1989). Stevenson and Iversen have conjugated polylysine to oligonucleotide analogs targeted to the splice donor and the 5'-end of the first exon of the tat gene. M. Stevenson, P. L. Iversen, J. Gen. Virol. 70:2673 (1989). Each of these publications have reported some degree of success in inhibiting some function of the HIV virus. While each of these references is distinct from the approach of the present invention, each supports the view that nucleotide therapeutics in HIV infection is rational and based upon sound scientific principles. In each of these references the approach has been to design antisense oligonucleotides complementary to some portion of the HIV mRNA. The present invention relates to oligonucleotides which mimic an RNA and bind to a protein, rather than oligonucleotides which bind to the HIV RNA.
Heretofore, there have been no suggestions in the art of methods or materials which could be useful for mimicking the secondary or tertiary structures of RNA in order to modulate the expression of genes or to treat disease. This is despite the long-felt need for methods of therapeutics and for methods of inhibiting gene expression which may be related to diseases or disease states in animals. Accordingly, there remains a long-felt need for therapeutic materials and methods, especially for viruses and retroviruses.